A large number of target molecules i.e. compounds and other materials e.g. peptides, vitamins, hormones, lipids, proteins, enzymes, membranes, cells and the like and compounds from e.g. a petroleum or coal source can be reacted, separated and/or purified using chromatographic techniques. Such techniques include ion-exchange chromatography, reverse phase chromatography, hydrophobic interaction chromatography, affinity chromatography and mixed mode chromatography. For purification of a target molecule, the techniques are based on characteristics of the target molecule such as solubility, charge, size, shape or affinity, which cause the target molecule to be retained due to inter-actions/reactions with the chromatographic resin.
The affinity chromatography techniques make use of a specific binding between the target molecule and certain support particles or conglomerates e.g. as those described in WO 92/00799. Thus, a solution containing the target macromolecule to be purified is passed through a column containing an insoluble support (also termed a matrix material or a resin). The molecules that do not exhibit appreciable affinity for the matrix material will pass through the column, whereas those molecules that recognise the matrix material, and react with it, will be retained. The specifically adsorbed target molecule can then be eluted by any one of a number of procedures, which will effect dissociation by e.g. altering the composition or the pH of the carrying medium.
To improve effectiveness of affinity chromatography the use of fluid bed reactor systems (including fluidised and expanded bed systems) has been developed (Chase, H. A. and Draeger, N. M. 1992. Affinity purification of proteins using expanded beds. Journal of Chromatography 597:129-145). These fluid bed systems make it possible to apply liquids such as fermentation broths containing, in addition to the target molecule, whole cells, broken or ruptured cells or other materials with the potential of interfering with the binding of the target molecule to the chromatographic resins. In more conventional systems such other materials could cause problems manifested by an increase in the pressure drop across the fluid bed and the formation of a plug of trapped solids at the inlet of the bed. Accordingly, in such conventional systems it is most often necessary to apply a purification step before loading the material onto the chromatographic column. The fluidised or expanded bed systems have overcome these problems. In these systems the bed of particles is fluidised or expanded in an unconstrained configuration by a flow of fluid (up-flow or down-flow). When a critical minimum liquid or gas velocity is exceeded, the bed particles start to expand and gaps occur between the adsorbent particles which will allow particles other than the target molecule to pass freely through the bed. The fluid bed systems have been further improved by making support particles (matrix materials or resins) available which effect binding of specific target molecules and subsequently effect desorption or elution of the target molecules, thereby retaining the target molecules in the fluid bed and reducing the risk that the target molecules will pass through the system by the flow of the carrying medium.
In the fluid bed systems a headspace adjacent to the bed particles is most often needed in order to eliminate the risk that the bed particles will agglomerate at the top of the column as a consequence of the flow of the carrying medium. Therefore, most fluid bed systems are equipped with a membrane near the outlet adjacent to a volume substantially void of the bed particles, i.e. a headspace, which membrane is impermeable to the bed particles but permits the carrying medium to pass through the system. Such a headspace is most often needed in order to make the fluidised and expanded bed systems work. However, in most situations the size of the headspace volume must be kept at a minimum as the target molecules to be purified will be diluted by the presence of excess medium. Accordingly, it is of vital importance to detect and control the dynamics of the fluidised or expanded system in order to control the headspace volume and at the same time expand the matrix material to a degree that secures optimal purification of the target molecule.
Methods for detection and subsequent control of the particle expansion in fluid bed systems are known. Currently used control systems include the use of ultrasonic reflection from the interface between the carrying medium substantially void of particles (headspace) and the bed particles. This is, however, an unreliable method as gas bubbles and solid particles trapped in the headspace volume may disturb the signals. The presence of gas bubbles will cause the signal to disappear and solid particles in the headspace volume will give rise to noise signals.
In U.S. Pat. No. 4,684,456, a method for control of bed expansion is described. The method comprises the use of sources of radiation and radiation detectors, which are fixed pair-wise along the chromatographic column at a first, second and a third level. The detectors detect an attenuated signal whenever the bed surface is above the level of the detector and its corresponding radiation source. Hence, the position of the bed surface can be detected only as lying within one of the intervals defined by the levels. Accordingly, the expansion of the bed particles is only controlled by an on/off signal. Such a signal does not allow for a dynamic control of the expansion, as there will be no information on how far the bed surface is from any of the levels, and accordingly no possibility of adjusting the flow velocity to this situation.
In an SU Inventors Certificate No. 1696886 it is recognised that the interface between the particle bed and the headspace may be inhomogeneous and thus difficult and unreliable to use for control of the system dynamics. A method is described which detects the headspace, an “inhomogeneous phase boundary”, in an expanded bed system. The method comprises providing a source of light and a sensor maintained at the same position in the fluid bed but not pointing towards each other. The sensor measures light from the source scattered by particles in the bed and thereby detects an average density coefficient of the fluidised particles. The method is based on performing repeated measurements under different flow conditions resulting in different volumes of the particle bed and consequently, different levels of the “inhomogeneous phase boundary” between the bed particles and the headspace. Since different volumes of the particle bed are related to different average density coefficient, a relationship between the flow velocity and the measured average density coefficient of the fluidised particles can be obtained. This relationship can subsequently be correlated to the pre-established position of the “inhomogeneous phase boundary” in the system. This approach has to be repeated for each individual system that is to be controlled and a great amount of time has to be spent on calibrating the system before use.
In the EP patent application EP 308027 A, a method is disclosed for controlling the suspension density of a particulate solids and gas mixture from a vessel, to yield a uniform and constant mass flow to a reactor. The method comprises the use of sources of radiation and radiation detectors opposite, which are fixed pair-wise. The detector determines the suspension density, which is compared to a pre-selected value. It appears that EP 308027 A does not reveal a method for density gradient detection or for dynamic control of an expansion, which implies that no disclosure is given as to a dynamical adjustment of the flow according to a determined value of the density.
In a French patent application FR 655053 A, a method is provided for controlling the mass flow to a catalysed cracking process of carbohydrates. The method comprises the use of sources of ionised radiation and radiation detectors to measure the suspension density. The suspension density determined by the detector is compared to a pre-selected value. It is obvious that FR 655053 A does not describe neither density gradient detection nor dynamic control of an expansion. Hence, this patent application provides no method for dynamically adjusting the flow according to the determined density.
Accordingly, the prior art discloses unreliable or cumbersome ways of controlling the expansion of a fluid bed reactor system. It is well known that the flow velocity through the fluid bed system is the determinant of the other dynamics of the system including the volume of the headspace. Therefore, a control system is needed which, in a fast and reliable manner, detects the dynamics of a fluidised or expanded bed and controls the system by continuously adjusting the flow velocity of the carrying medium.
Such a control system could benefit from a transfer-function between a sensor activating signal and a sensor-output signal characterised as a continuous monotone function and the response time would be much faster and the system more accurate than known systems. Furthermore, such a sensor system will generate a much faster and more precise signal if the distance from the interface between the headspace medium and the mixture of medium and bed particles is based on measurements of a particular particle density gradient in the particle bed and thus, not simply relying on an on/off signal.